1. Field of the Invention
The present invention relates to measuring cells adapted for micro-assays, notably for selective micro-assays of constituents in biological fluids, and comprising membrane electrodes. The invention also relates to an improved electrode carrying a membrane.
2. Description of the Prior Art
From the original studies of S. Updike and G. Kicks (Nature, Lond.--1967, 214, 986) is is known to be possible to assay selectively a substance in a heterogeneous biological fluid by using a conventional polarographic electrode (with a partial pressure of oxygen pO.sub.2, for example) coated with a membrane bearing the specific enzyme of the substance to be assayed. Updike and Kicks have thus determined glucose with a pO.sub.2 electrode coated with a polyacrylamide gel bearing glucoseoxydase included in the gel.
Since these initial studies, other investigations have been carried out, which have borne on the nature of the membranes, on the types of enzymes applicable and on the possibilities of utilizing different electrodes. In this respect, should be mentioned:
the article of G. G. Guilbault and J. Montalvo (J. Amer. Chem. Soc., 1969, 91, 2164) relating to determination of urea by a cation electrode equipped with an inclusion of urease in a polyacrylamide gel;
the article of G. G. Guilbault and E. Hrabankova (Analyt. Chem., 1970, 42,1779) relating to determination of total aminoacids by a pO.sub.2 electrode equipped with an oxidase aminoacid inclusion and polyacrylamide;
French Pat. No. 1,604,982, as well as the first and second Certificates of Addition associated therewith, bearing respectively Nos. 69-01451 and 69-07897, wherein various modifications are described of a method for the preparation of enzymatic membranes having very much improved properties with respect to inclusions in the polyacrylamide; and
the article of Berjonneau A. M., Thomas D. Broun G. (Path. Biol., 1974, 497, 502) entitled "Specific Assay of Aminoacids by pCO.sub.2 Electrodes Equiped with Decarboxylase Membranes by Protein Coreticulation".
In addition, work carried out since 1969 by teams including the last-mentioned researchers have enabled full definition of the conditions of preparation of enzymatic membranes adapted for use with standard polarographic electrodes of various types, such as notably pO.sub.2 and pCO.sub.2 (see on this subject the article of Calvot C. et al in FEBS Letters, Vol. 59 No. 2, November 1975, pages 258-262 and the article of Cordonnier M. et al. in FEBS Letters, Vol. 59, No. 2, November 1975, pages 263-267).
Up to the present, these membranes have been used with commercial electrodes, for example electrodes of the type known under the tradename Radiometer; in such cases, the membrane is mounted on the sheath of the electrode by means of a toric seal of rubber. This method is delicate in assembly and, in addition, it causes mechanical stresses and irregular tension at the level of the membrane itself, so that it is impossible to avoid a notable percentage of torn membranes, as well as poor durability. Moreover, the size of the outer body of the electrode is then such that it is not possible to use an electrode of this design on microcells utilizing an overall volume of the analysis compartment of only 20 microliters.
In the article in Analytical Chemistry, Vol. 43, No. 11, September 1971, by R. A. Lienado et al, there is only described a measuring electrode bearing a membrane held in place by a cap of Plexiglas which includes a single cavity, without orifices for communication with the exterior, whilst an annular seal of rubber must be present to ensure fluid-tightness which is even then doubtful.
In the U.S. Pat. No. 3,151,052, an electrochemical cell of old design not including an enzymatic membrane, is described, and no suggestion is contained therein for utilizing such a membrane therewith.
It is an object of the present invention to provide an improved measuring cell for micro-assays comprising membrane electrodes, e.g. enzymatic membrane electrodes.
Other objects and advantages of the present invention will emerge from the description which follows.